Backgroud: Myocardial infarction (MI) is one of the leading causes of global death. Dendritic cell-derived exosomes (DEXs) provide us with the possibility of improving cardiac function after MI but are limited by low retention times and short-lived therapeutic effects. In this study, we developed a novel drug delivery system incorporating alginate hydrogel that continuously releases DEXs and investigated the mechanisms underlying the action of DEXs in the improvement of cardiac function after MI. Results: We incorporated DEXs with alginate hydrogel (DEXs-Gel) and investigated controlled released ability and rheology, and found that DEXs-Gel release DEXs in a sustainable mammer and prolonged the retention time of DEXs but had no detrimental effects on the migration in vivo. Then DEXs-Gel was applicated in the MI model mice, we found that DEXs-Gel siginificantly enhanced the therapeutic effects of DEXs with regards to improving cardiac function after MI. Flow cytometry and immunofluorescence staining revealed that DEXs significantly upregulated the infiltration of Treg cells and M2 macrophages into the border zoom after MI, and DEXs activated regulatory T (Treg) cells and shifted macrophages to reparative M2 macrophages, both in vitro and in vivo. Conclusion: Our novel delivery method provides an innovative tool for enhancing the therapeutic effects of DEXs after MI. Further analysis revealed that DEXs exert effect by activating Treg cells and by modifying the polarization of macrophages. Graphic

Background: Few study has been done to evaluate the stability and superiority of normalizers for serum microRNA (miRNA) study in cardiovascular disease. Therefore, the aim of this study is to assess the suitability of several common normalizers (miR-16, SNOU6, 5S, miR-19b, miR-24, miR-15b, let 7i) in cardiovascular disease. Methods: We evaluated the stability of the seven circulating miRNAs as reference genes in the blood samples from patients with cardiovascular disease [heart failure (HF) and hypertension] and healthy people. Stability was quantified by combining BestKeeper, NormFinder and comparative delta Cq analysis. Results: A total of 62 subjects were included in this study, of which 25 patients were with HF, 10 patients were with hypertension, and 27 were healthy people. The analysis from both BestKeeper and comparative delta ct analysis demonstrated that let-7i and miR-16 showed the best performance [the standard deviations (SD) in BestKeeper for let-7i and miR-16 were 0.60 and 0.72, and the mean SD in comparative delta ct analysis for let-7i and miR-16 were 1.79 and 1.82, respectively], while SNOU6 and 5S had the highest variability. In NormFinder analysis, miR-15 show best stability (ρ=0.029), followed by miR-19b (ρ=0.037), let-7i (ρ=0.064), SNOU6 (ρ=0.064), 5S (ρ=0.064), miR-16 (ρ=0.064), while miR-24 (ρ=0.075) showed worst stability. Conclusions: This study pointed out that in the serum studies focused on cardiovascular disease, let-7i and miR-16 had the best performance, while SNOU6 and 5S were not suitable as reference gene. This study indicate that the selection of an optimal reference genes is important to get an accurate result in serum miRNA studies, the findings are of clinical significance to guide the further miRNA studies or tests.

Dendritic cells (DCs) play key roles in promoting wound healing after myocardial infarction (MI). Our previous studies have shown that exosomes derived from DCs (DEXs) could migrate to lymphoid tissue and improve cardiac function post-MI by activating CD4⁺ T cells; however, the mechanism of DEXs’ migration to lymphoid tissue and the improvement of cardiac function are still unknown. In our study, we found that CCR7 expression significantly increased in MI-DEXs compared with control-DEXs; meanwhile, CCL19 and CCL21, the ligands of CCR7, significantly increased in the serum of MI-model mice. Subsequently, we overexpressed and knocked down CCR7 in MI-DEXs and found that overexpressed CCR7 enhanced the migration of MI-DEXs to the spleen; however, CCR7 knockdown attenuated MI-DEXs’ migration according to near-IR fluorescence imaging. Furthermore, overexpressed CCR7 in MI-DEXs enhanced the MI-DEXs’ improvement of cardiac function after MI; however, CCR7-knockdown MI-DEXs attenuated this improvement. In addition, after DEXs’ migration to the spleen, MI-DEXs activated CD4⁺ T cells and induced the expression of IL-4 and IL-10, which were significantly increased in the MI-DEX group compared with the control group. In conclusion, CCR7 could mediate DEXs’ migration to the spleen and improve cardiac function after MI, and we found that the mechanism was partly via activation of CD4⁺ T cells and secretion of IL-4 and IL-10. Our study presented an innovative method for improving cardiac function by enhancing the migration ability of MI-DEXs after MI, while CCR7 could be a potential candidate for MI-DEX bioengineering to enhance migration.

Revascularization of coronary chronic total occlusion (CTO) still remains controversial. The factors that impact collateral circulation and myocardial perfusion are of interest. Circular RNA (circRNA) has been shown to regulate the process of angiogenesis. However, the effects of circ-membrane-bound O-acyltransferase domain containing 2 (circ-MBOAT2) on angiogenesis in patients with CTO were unclear. In this study, we evaluated circulating circRNAs and miRNAs in patients with CTO and stable coronary artery disease using high-throughput sequencing. Another cohort of patients were selected to verify the expressions of circ-MBOAT2 and miR-495. The role and mechanism of circ-MBOAT2 in the process of angiogenesis were explored through in vitro and vivo studies. Finally, we came back to a clinical perspective and investigated whether circ-MBOAT2 and miR-495 were associated with the improvement of myocardial perfusion evaluated by single-photon emission computed tomography (SPECT). We found that the expression of circ-MBOAT2 was significantly up-regulated while miR-495 was significantly down-regulated in patients with CTO. The expression of circ-MBOAT2 was negatively correlated with miR-495 in patients with CTO. In an in vitro study, we found that circ-MBOAT2 promoted tube formation and cell migration via the miR-495/NOTCH1 axis in endothelial cells. In an in vivo study, we showed that the inhibition of miR-495 caused the increase in collateral formation in mice after hindlimb ischemia. In a human study, we showed the expressions of circ-MBOAT2 and miR-495 were associated with myocardial perfusion improvement after revascularization of CTO. In conclusion, circ-MBOAT2 regulates angiogenesis via the miR-495/NOTCH1 axis and associates with myocardial perfusion in patients with CTO. Our findings suggest that circ-MBOAT2 and miR-495 may be potential therapeutic targets and prognostic factors for patients with CTO.

Background: Coronary chronic total occlusion (CTO) disease is common and its specific characteristic is collateral formation. The Integrated analysis of angiogenesis related lncRNA-miRNA-mRNA network remains unclear and might provide target for future studies.Methods: A total of five coronary artery disease (control group) and five CTO (CTO group) patients were selected for deep RNA and miRNA sequencing. The expression profiles of lncRNAs, mRNAs circRNA and miRNAs were obtained. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were then performed. The expression of a 14q32 miRNA gene cluster, including miRNA-494, miRNA-495 and miRNA-329, were selected to be determined in another larger patient cohort. Analysis of the lncRNA-miRNA495-mRNA network was constructed to find potential targets for future studies.Results: A total of 871 lncRNAs, 1,080 mRNAs, 138 circRNAs and 56 miRNAs were determined as differentially expressed (DE) in CTO patients compared with control patients. GO and KEGG analyses revealed that the top terms included MAPK signaling pathway, HIF-1 signaling pathway, EGFR tyrosine kinase inhibitor resistance, embryonic organ development, wound healing, MAPK signaling pathway and JAK-STAT signaling pathway, which are related to angiogenesis. The expression of miRNA-494, miRNA-495 and miRNA-329 were all significantly down-regulated in CTO patients and they were confirmed to be down-regulated in another cohort of 68 patients. Then we divided the CTO patients into two groups according to CC grade (poor CC group, CC = 0 or one; good CC group, CC = 2). MiRNA-494, miRNA-495 and miRNA-329 were found to be down-regulated in good CC group compared with poor CC group. Analysis of the lncRNA-miRNA495-mRNA network showed 3 DE lncRNA sponges (NONHSAG008675, NONHSAG020957 and NONHSAG010989), 4 DE lncRNA targets (NONHSAT079547.2, NONHSAT081776.2, NONHSAT148555.1 and NONHSAT150928.1) and 2 DE mRNA targets (RAD54L2 and ZC3H4) of miRNA495.Conclusion: This study revealed that the lncRNA-miRNA-mRNA network might play a critical role in angiogenesis in CTO patients.